Structural analysis of substrate binding by the TatBC component of the twin-arginine protein transport system.
Structural analysis of substrate binding by the TatBC component of the twin-arginine protein transport system.
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双精氨酸蛋白转运系统 TatBC 成分结合底物的结构分析。
DOI：
10.1073/pnas.0901566106
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发表时间：
2009
影响因子：
11.1
通讯作者：
Tarry MJ
中科院分区：
文献类型：
--
作者：
Tarry MJ
达特系统将折叠的蛋白质运输穿过细菌细胞质膜和植物叶绿体的类囊体膜。在大肠杆菌底物蛋白最初结合到完整的膜TatBC复合物，然后招募蛋白质TatA的影响易位。在不存在TatA的情况下，TatBC和底物蛋白SufI的过量产生导致TatBC-SufI复合物的积累，该复合物可以使用底物上的亲和标签进行纯化。通过单粒子电子显微镜和随机锥形倾斜重建获得了TatBC-SufI复合物和未配体的TatBC的三维结构。结构的比较表明，底物分子结合在TatBC复合物的外周上，并且底物结合导致复合物的TatBC部分的直径显著减小。尽管TatBC复合物含有多个拷贝的信号肽结合TatC原聚体，但纯化的TatBC-SufI复合物仅含有1或2个SufI分子。当TatBC-SufI复合物中存在2种底物时，它们在相邻位点结合。这些观察结果意味着，只有某些TatC原聚体内的复杂与基板相互作用，或有一个负协同性的基板结合。类似的TatBC-底物复合物可以通过替代的体外重构方法和使用不同的底物蛋白质来产生。
The Tat system transports folded proteins across the bacterial cytoplasmic membrane and the thylakoid membrane of plant chloroplasts. InEscherichia colisubstrate proteins initially bind to the integral membrane TatBC complex which then recruits the protein TatA to effect translocation. Overproduction of TatBC and the substrate protein SufI in the absence of TatA led to the accumulation of TatBC-SufI complexes that could be purified using an affinity tag on the substrate. Three-dimensional structures of the TatBC-SufI complexes and unliganded TatBC were obtained by single-particle electron microscopy and random conical tilt reconstruction. Comparison of the structures shows that substrate molecules bind on the periphery of the TatBC complex and that substrate binding causes a significant reduction in diameter of the TatBC part of the complex. Although the TatBC complex contains multiple copies of the signal peptide-binding TatC protomer, purified TatBC-SufI complexes contain only 1 or 2 SufI molecules. Where 2 substrates are present in the TatBC-SufI complex, they are bound at adjacent sites. These observations imply that only certain TatC protomers within the complex interact with substrate or that there is a negative cooperativity of substrate binding. Similar TatBC-substrate complexes can be generated by an alternative in vitro reconstitution method and using a different substrate protein.